scholarly journals Phloretin Suppresses Bone Morphogenetic Protein-2-Induced Osteoblastogenesis and Mineralization via Inhibition of Phosphatidylinositol 3-kinases/Akt Pathway

2019 ◽  
Vol 20 (10) ◽  
pp. 2481 ◽  
Author(s):  
Ayumu Takeno ◽  
Ippei Kanazawa ◽  
Ken-ichiro Tanaka ◽  
Masakazu Notsu ◽  
Toshitsugu Sugimoto
Keyword(s):  
Osteoblast Differentiation ◽  
Glucose Transporter ◽  
Binding Protein ◽  
Bone Morphogenetic Protein 2 ◽  
Akt Pathway ◽  
Akt Inhibitor ◽  
Differentiation Markers ◽  
Uptake Inhibition ◽  
Fatty Acid Binding ◽  
Phosphatidylinositol 3

Phloretin has pleiotropic effects, including glucose transporter (GLUT) inhibition. We previously showed that phloretin promoted adipogenesis of bone marrow stromal cell (BMSC) line ST2 independently of GLUT1 inhibition. This study investigated the effect of phloretin on osteoblastogenesis of ST2 cells and osteoblastic MC3T3-E1 cells. Treatment with 10 to 100 µM phloretin suppressed mineralization and expression of osteoblast differentiation markers, such as alkaline phosphatase (ALP), osteocalcin (OCN), type 1 collagen, runt-related transcription factor 2 (Runx2), and osterix (Osx), while increased adipogenic markers, peroxisome proliferator-activated receptor γ (PPARγ), CCAAT/enhancer-binding protein α (C/EBPα), fatty acid-binding protein 4, and adiponectin. Phloretin also inhibited mineralization and decreased osteoblast differentiation markers of MC3T3-E1 cells. Phloretin suppressed phosphorylation of Akt in ST2 cells. In addition, treatment with a phosphatidylinositol 3-kinase (PI3K)/Akt inhibitor, LY294002, suppressed the mineralization and the expression of osteoblast differentiation markers other than ALP. GLUT1 silencing by siRNA did not affect mineralization, although it decreased the expression of OCN and increased the expression of ALP, Runx2, and Osx. The effects of GLUT1 silencing on osteoblast differentiation markers and mineralization were inconsistent with those of phloretin. Taken together, these findings suggest that phloretin suppressed osteoblastogenesis of ST2 and MC3T3-E1 cells by inhibiting the PI3K/Akt pathway, suggesting that the effects of phloretin may not be associated with glucose uptake inhibition.

scholarly journals Phosphatidylinositol 3-Kinase/Akt Pathway Targets Acetylation of Smad3 through Smad3/CREB-binding Protein Interaction

2009 ◽  
Vol 284 (36) ◽  
pp. 23912-23924 ◽  
Author(s):  
Lassad Oussaief ◽  
Aurélie Hippocrate ◽  
Vanessa Ramirez ◽  
Aurore Rampanou ◽  
Wei Zhang ◽  
...  
Keyword(s):  
Protein Interaction ◽  
Binding Protein ◽  
Akt Pathway ◽  
Creb Binding Protein ◽  
Phosphatidylinositol 3 Kinase ◽  
Phosphatidylinositol 3

scholarly journals Food for Bone: Evidence for a Role for Delta-Tocotrienol in the Physiological Control of Osteoblast Migration

2020 ◽  
Vol 21 (13) ◽  
pp. 4661 ◽  
Author(s):  
Lavinia Casati ◽  
Francesca Pagani ◽  
Roberto Maggi ◽  
Francesco Ferrucci ◽  
Valeria Sibilia
Keyword(s):  
Wound Healing ◽  
Osteoblast Differentiation ◽  
Target Genes ◽  
Elaeis Guineensis ◽  
Signalling Pathway ◽  
Bone Morphogenetic Protein 2 ◽  
Luciferase Assay ◽  
Boyden Chamber ◽  
Differentiation Markers ◽  
Osteoblast Migration

Bone remodeling and repair require osteogenic cells to reach the sites that need to be rebuilt, indicating that stimulation of osteoblast migration could be a promising osteoanabolic strategy. We showed that purified δ-tocotrienol (δ-TT, 10 μg/mL), isolated from commercial palm oil (Elaeis guineensis) fraction, stimulates the migration of both MC3T3-E1 osteoblast-like cells and primary human bone marrow mesenchymal stem cells (BMSC) as detected by wound healing assay or Boyden chamber assay respectively. The ability of δ-TT to promote MC3T3-E1 cells migration is dependent on Akt phosphorylation detected by Western blotting and involves Wnt/β-catenin signalling pathway activation. In fact, δ-TT increased β-catenin transcriptional activity, measured using a Nano luciferase assay and pretreatment with procaine (2 µM), an inhibitor of the Wnt/β-catenin signalling pathway, reducing the wound healing activity of δ-TT on MC3T3-E1 cells. Moreover, δ-TT treatment increased the expression of β-catenin specific target genes, such as Osteocalcin and Bone Morphogenetic Protein-2, involved in osteoblast differentiation and migration, and increased alkaline phosphatase and collagen content, osteoblast differentiation markers. The ability of δ-TT to enhance the recruitment of BMSC, and to promote MC3T3-E1 differentiation and migratory behavior, indicates that δ-TT could be considered a promising natural anabolic compound.

scholarly journals Statin-induced Ras Activation Integrates the Phosphatidylinositol 3-Kinase Signal to Akt and MAPK for Bone Morphogenetic Protein-2 Expression in Osteoblast Differentiation

2006 ◽  
Vol 282 (7) ◽  
pp. 4983-4993 ◽  
Author(s):  
Nandini Ghosh-Choudhury ◽  
Chandi Charan Mandal ◽  
Goutam Ghosh Choudhury
Keyword(s):  
Bone Morphogenetic Protein ◽  
Osteoblast Differentiation ◽  
Dominant Negative ◽  
Bone Morphogenetic Protein 2 ◽  
Regulatory Subunit ◽  
Type I ◽  
Dependent Manner ◽  
Phosphatidylinositol 3 Kinase ◽  
Morphogenetic Protein ◽  
Phosphatidylinositol 3

Lovastatin promotes osteoblast differentiation by increasing bone morphogenetic protein-2 (BMP-2) expression. We demonstrate that lovastatin stimulates tyrosine phosphorylation of the p85 regulatory subunit of phosphatidylinositol 3-kinase (PI3K), leading to an increase in its kinase activity in osteoblast cells. Inhibition of PI3K ameliorated expression of the osteogenic markers alkaline phosphatase, type I collagen, osteopontin, and BMP-2. Expression of dominant-negative PI3K and PTEN, an inhibitor of PI3K signaling, significantly attenuated lovastatin-induced transcription of BMP-2. Akt kinase was also activated in a PI3K-dependent manner. However, our data suggest involvement of an additional signaling pathway. Lovastatin-induced Erk1/2 activity contributed to BMP-2 transcription. Inhibition of PI3K abrogated Erk1/2 activity in response to lovastatin, indicating the presence of a signal relay between them. We provide, as a mechanism of this cross-talk, the first evidence that lovastatin stimulates rapid activation of Ras, which associates with and activates PI3K in the plasma membrane, which in turn regulates Akt and Erk1/2 to induce BMP-2 expression for osteoblast differentiation.

scholarly journals Intestinal maturation in mice lacking CCAAT/enhancer-binding protein α (C/EPBα)

1998 ◽  
Vol 330 (3) ◽  
pp. 1165-1171 ◽  
Author(s):  
J. Thomas OESTERREICHER ◽  
L. Lucy LEEPER ◽  
J. Milton FINEGOLD ◽  
J. Gretchen DARLINGTON ◽  
J. Susan HENNING
Keyword(s):  
Glucose Transporter ◽  
Binding Protein ◽  
Fatty Acid Binding Proteins ◽  
Mrna Levels ◽  
Fatty Acid Binding ◽  
Proliferative Zone ◽  
Glucose Transporter 2 ◽  
Enhancer Binding Protein ◽  
Ccaat Enhancer Binding Protein ◽  
Morphological Maturation

In rodents, there is a surge of intestinal expression of CCAAT/enhancer-binding protein α (C/EBPα) in the late fetal phase just before morphological maturation and the onset of expression of numerous epithelial genes. To investigate directly the hypothesis that C/EBPα plays a causal role in the latter phenomena, we have assessed both structural and functional maturation in neonatal intestine from C/EBPα-null mice and their littermates. No effects of C/EBPα genotype were observed on mucosal architecture or on the size of the proliferative zone in the intestinal crypts. Likewise, the mRNA levels for the glucose transporter 2 (GLUT2), intestinal and liver fatty acid-binding proteins, and apolipoprotein A-IV in newborn intestine were similar in all genotypes. Paradoxically, Na+/glucose co-transporter (SGLT1), lactase phlorizin-hydrolase and apolipoprotein B mRNAs were more abundant in the C/EBPα-deficient animals. In wild-type intestines, C/EBPβ and C/EBPΔ mRNAs were detectable throughout the late fetal period and increased toward term in parallel with C/EBPα mRNA. In newborn intestine, there was no compensatory up-regulation of these isoforms in the C/EBPα-deficient mice. We conclude that C/EBPα has no essential role in morphological maturation of the intestine, the pattern of proliferation of the epithelium, or the onset of expression of this cluster of epithelial mRNAs. However, since other C/EBP isoforms are present in the developing intestine, it is possible that there is a generic requirement for a member of the C/EBP family.

scholarly journals Phosphatidylinositol 3 Kinase/Akt Signal Relay Cooperates with Smad in Bone Morphogenetic Protein-2-Induced Colony Stimulating Factor-1 (CSF-1) Expression and Osteoclast Differentiation

Endocrinology ◽  
2009 ◽  
Vol 150 (11) ◽  
pp. 4989-4998 ◽  
Author(s):  
Chandi C. Mandal ◽  
Goutam Ghosh Choudhury ◽  
Nandini Ghosh-Choudhury
Keyword(s):  
Binding Protein ◽  
Osteoclast Differentiation ◽  
Osteoclast Formation ◽  
Bone Morphogenetic Protein 2 ◽  
Colony Stimulating Factor ◽  
Phosphatidylinositol 3 Kinase ◽  
Osteoblast Activity ◽  
Stimulating Factor ◽  
Phosphatidylinositol 3

Murine spleen cells produce mature osteoclasts when cocultured with osteoblastic cells. Colony-stimulating factor (CSF)-1 is the growth factor required for differentiating the monocyte-macrophage precursor cells into preosteoclasts. Bone morphogenic protein (BMP) signaling in osteoblasts regulates bone mass in mice, suggesting a role of BMP in osteoclastogenesis along with osteoblast activity. The intracellular signal transduction cross talk regulating the osteoblastic production of CSF-1 as a mechanism of BMP-induced osteoclastogenesis is described in this report. We have recently described the involvement of Smad 1/5 in BMP-2-induced CSF-1 expression and osteoclast formation. In this study, using the pharmacological inhibitors and the adenovirus (Ad) vectors expressing dominant-negative (DN) phosphatidylinositol 3 kinase (PI3K), the PI3K-signaling inhibitor, phosphatase and tensin homolog deleted in chromosome 10 (PTEN) or DN Akt kinase in the in vitro coculture assay, we show an essential role of the lipid kinase cascade in BMP-2-mediated multinucleated osteoclast formation and CSF-1 mRNA expression, transcription, and secretion. Inhibition of PI3K/Akt signaling blocked the binding of Smads 1/5 to the CSF-1 BMP-responsive element present in the CSF-1 promoter, resulting in attenuation of Smad-dependent CSF-1 transcription. Furthermore, PI3K inhibition and DN Akt prevented association of the transcriptional coactivator, CREB (cAMP response element binding protein) binding protein (CBP), with Smads 1/5. Together, these data for the first time demonstrate that PI3K-dependent Akt activation regulates BMP-2-induced CSF-1 expression and provides a mechanism for osteoblastic cell-assisted osteoclast differentiation.

C2C12 myoblast/osteoblast transdifferentiation steps enhanced by epigenetic inhibition of BMP2 endocytosis

2002 ◽  
Vol 283 (1) ◽  
pp. C235-C243 ◽  
Author(s):  
Cyril Rauch ◽  
Anne-Christine Brunet ◽  
Julie Deleule ◽  
Emmanuel Farge
Keyword(s):  
Osteoblast Differentiation ◽  
Nuclear Translocation ◽  
Bone Morphogenetic Protein 2 ◽  
C2c12 Myoblast ◽  
Differentiation Markers ◽  
Signaling Protein ◽  
Mechanical Deformations ◽  
Morphogenetic Protein ◽  
Cell Genome ◽  
Differentiation Marker

We investigated the modulation of critical transcriptional steps of C2C12myoblast/osteoblast transdifferentiation triggered by the bone morphogenetic protein 2 (BMP2) signaling protein, in response to epigenetic inhibition of the endocytotic internalization of exogenous BMP2. BMP2 endocytosis was inhibited chemically with polyethylene glycol-50 (PEG-Chol) and cyclodextrin and mechanically by mild hyposmotic treatment. BMP2-dependent nuclear translocation of the mother against Dpp (Smad1) transcription factor was ten times faster if BMP2 endocytosis was inhibited. Smad1-dependent expression of the JunB gene, the first transcriptional step in myoblast dedifferentiation, was increased by a factor of three to four. JunB-dependent levels of myogenin repression, one of the critical markers of terminal myoblastic differentiation, was amplified by a factor of three. Smad1-dependent levels of alkaline phosphatase expression, one of the C2C12 osteoblast differentiation markers, were 3.5 to 5 times higher. The same behavior was observed for osteopontin, the other C2C12 osteoblast differentiation marker. These results suggest that the cell genome could “sense” tissue mechanical deformations by mechanical inhibition of signaling protein endocytosis, thereby translating mechanical strains into transcription events involved in cell differentiation.

scholarly journals Activation of p38 Mitogen-Activated Protein Kinase Is Required for Osteoblast Differentiation

Endocrinology ◽  
2003 ◽  
Vol 144 (5) ◽  
pp. 2068-2074 ◽  
Author(s):  
Yuanyu Hu ◽  
Emily Chan ◽  
Sherry X. Wang ◽  
Baojie Li
Keyword(s):  
Bone Marrow ◽  
P38 Mapk ◽  
Osteoblast Differentiation ◽  
Specific Inhibitor ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Bone Morphogenetic Protein 2 ◽  
Differentiation Markers ◽  
Mineral Deposition ◽  
Calvarial Osteoblast

p38 MAPK is a conserved subfamily of MAPKs involved in inflammatory response, stress response, cell growth and survival, as well as differentiation of a variety of cell types. In this report we demonstrated that p38 MAPK played an important role in osteoblast differentiation using primary calvarial osteoblast, bone marrow osteoprecursor culture, and a murine cell line, MC3T3-E1. We found that p38 MAPK was activated as calvarial osteoblast differentiates along with extracellular signal-regulated kinases (ERKs). When p38 MAPK is inhibited with a specific inhibitor, the expression of differentiation markers, such as alkaline phosphatase and mineral deposition, were significantly reduced. MC3T3-E1 cells expressing dominant negative p38 MAPK also displayed signs of delay in ALP and mineral deposition. Differentiation of the bone marrow osteoprecursors was also impeded by the p38 MAPK inhibitor, justified by the same markers. Yet the inhibitory effects observed in calvarial osteoblasts and bone marrow osteoprogenitor cells could be partially prevailed by bone morphogenetic protein-2. Inhibition of ERKs with a specific drug did not significantly affect osteoblast differentiation even though ERK1/2 were also activated during osteoblast differentiation. These results taken together indicate that p38 MAPK, but not ERKs, is necessary for osteoblast differentiation.

scholarly journals The Hormonal Response of Estrogen Receptor β Is Decreased by the Phosphatidylinositol 3-Kinase/Akt Pathway via a Phosphorylation-dependent Release of CREB-binding Protein

2006 ◽  
Vol 282 (7) ◽  
pp. 4830-4840 ◽  
Author(s):  
Mélanie Sanchez ◽  
Karine Sauvé ◽  
Nathalie Picard ◽  
André Tremblay
Keyword(s):  
Target Gene ◽  
Binding Protein ◽  
Intrinsic Activity ◽  
Akt Pathway ◽  
Creb Binding Protein ◽  
Transcriptional Coactivator ◽  
Phosphatidylinositol 3 Kinase ◽  
Hormonal Response ◽  
Reduced Activity ◽  
Phosphatidylinositol 3

The hormonal response of estrogen receptors (ER) α and ERβ is controlled by a number of cofactors, including the general transcriptional coactivator CREB-binding protein (CBP). Growing evidence suggests that specific kinase signaling events also modulate the formation and activity of the ER coactivation complex. Here we show that ERβ activity and target gene expression are decreased upon activation of ErbB2/ErbB3 receptors despite the presence of CBP. This inhibition of ERβ involved activation of the phosphatidylinositol 3-kinase/Akt pathway, abrogating the potential of CBP to facilitate ERβ response to estrogen. Such reduced activity was associated with an impaired ability of ERβ to recruit CBP upon activation of Akt. Mutation of serine 255, an Akt consensus site contained in the hinge region of ERβ, prevented the release of CBP and rendered ERβ transcriptionally more responsive to CBP coactivation, suggesting that Ser-255 may serve as a regulatory site to restrain ERβ activity in Akt-activated cells. In contrast, we found that CBP intrinsic activity was increased by Akt through threonine 1872, a consensus site for Akt in the cysteine- and histidine-rich 3 domain of CBP, indicating that such enhanced transcriptional potential of CBP did not serve to activate ERβ. Interestingly, nuclear receptors sharing a conserved Akt consensus site with ERβ also exhibit a reduced ability to be coactivated by CBP, whereas others missing that site were able to benefit from the activation of CBP by Akt. These results therefore outline a regulatory mechanism by which the phosphatidylinositol 3-kinase/Akt pathway may discriminate nuclear receptor response through coactivator transcriptional competence.

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